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The Centers
for Disease Control and Prevention (CDC) is working with local and state
health departments and public health laboratories to test specimens from
humans and animals with suspected monkeypox infection. This document provides
interim recommendations for the safe collection of human clinical specimens
for monkeypox testing.
Reporting
of Suspect Human Monkeypox Cases
Report possible
human cases of monkeypox to your local hospital epidemiologist and/or
infection control personnel, who will contact your state health department.
The state health department will contact the Centers for Disease Control
and Prevention (CDC), if appropriate. Consultation with the state
epidemiologist and state
health laboratory is necessary before sending specimens to CDC. After
CDC has been contacted for approval to send the specimens, the samples
should be collected in the manner outlined below and placed in an appropriate
biosafe shipping container.
Protection
of Personnel
Specimen
collection personnel. Personnel who collect specimens should
use personal protective equipment (PPE) in accordance with recommendations
for Standard, Contact, and Airborne Precautions. Current recommendations
for smallpox vaccination
of health-care workers also should be considered.
Laboratory
personnel. For guidance regarding the use of smallpox vaccine
for laboratory personnel who potentially might handle specimens from patients
with monkeypox, see Interim
Biosafety Guidelines for Laboratory Personnel.
Sharps
injury prevention. When possible, use plastic rather than glass
materials for specimen collection and processing (e.g., blood and capillary
tubes, bottles, slides) and devices with sharps injury prevention features.
Dispose all sharps in a sharps container immediately after use.
Processing
of routine clinical specimens. For information about laboratory
precautions when processing routine clinical specimens, see Interim
Biosafety Guidelines for Laboratory Personnel.
Collection
of Specimens for Monkeypox Diagnosis
This
section provides guidance for the collection of various clinical specimens
during the rash phases of monkeypox as well as the prodromal and convalescent
stages. Samples should be collected in the manner specified below
and placed in an appropriate biosafe shipping container.
I.
Specimen Collection Supply List
Some or all
of the following materials will be required for specimen collection from
each patient:
Blood
collection
- 5- or
10-cc syringe with 18- or 20-gauge needle (pediatrics: smaller syringe
and smaller gauge needle may be necessary)
- 1 vacutainer
holder
- 2 vacutainer
needles (20 x 1.5 in.)
- 1 10-cc
marble-topped vacutainer tube, or 1 10-cc yellow-topped serum separator
tube for serum collection (plastic tube preferable)
- 1 5-cc
purple-topped tube (potassium EDTA anticoagulant) for whole blood buffy
coat collection for viral isolation (plastic tube preferable)
- Styrofoam
tube protectors
Lesion
collection: macular, papular, vesicular, pustular,scabs
- Disposable
scalpel (1) with No. 10 blade
- Sterile
26-gauge needles (several)
- Punch
biopsy kit; 2.5- (pediatric), 3.5- or 4-mm
- 10% buffered
formalin
- Needle
driver
- Suture
- Suture
removal kit
- Sterile
dry polyester or Dacron swabs (e.g., Catch-All sample collection swabs,
catalog no. QEC091H from Epicentre in Madison, Wisconsin (Epicentre)
(4-8)
- Clean
plastic or glass microscope slides (4)
- Plastic
single-slide holders (4)
- Formvar/carbon-coated
mesh electron microscopy grids
(2-4), catalog no. FCF400-Cu; sterilize under a UV light for 10
minutes prior to use
- Electron
microscopy quality forceps, catalog
no. 72750-F
- Electron
microscopy grid box, catalog
no.71150
- Sterile
screw-capped plastic vials (1.5 to 2.0 ml) (Sarstedt with O-ring)
- Parafilm
Oropharyngeal
specimen collection
- Polyester
or Dacron swabs (e.g., Catch-All sample collection swabs, catalog no.
QEC091H from Epicentre in Madison, Wisconsin (Epicentre)
Other
- Sharps
container
- Disposable
biohazard bags
Disclaimer:
Names of vendors or manufacturers are provided as examples of suitable
product sources; inclusion does not imply endorsement by the Centers for
Disease Control and Prevention, Department of Health and Human Services.
II.
Specimen Types and Labeling Procedure
Suitable
specimens for virologic tests are:
- Vesicular
or pustular tissue and fluid
- Scabs
- Biopsy
tissues
- Throat
swabs and whole blood
Suitable
specimen for serologic tests are:
Specimen
Labeling: Label all tubes, vials, microscope
slides, and EM grid holders with the following:
- Patient
name
- Date and
time of collection
- Source
of specimen (vesicle, pustule, or scab)
- Date of
birth of patient (for cross-referencing of specimens)
- Name
or initials of person collecting specimen
- If patient
is hospitalized, include hospital and identification numbers (e.g.,
medical record number, surgical pathology number)
- State
ID number or CDC monkeypox ID number.
III.
Pre-collection Procedures
IV.
Stepwise Collection Procedures for Patients with Acute Symptoms of Monkeypox
Rash lesions:
Macular*, papular*, vesicular, or pustular lesions
- Sanitize
skin with an alcohol wide, allow to dry.
- Use scalpel
(or a sterile 26-gauge needle) to open, and remove, the top of the vesicle
or pustule. Do not send the scalpel or sharp. Dispose of in appropriate
biohazard container and dispense.
- Place
the skin of the vesicle top into a 1.5- to 2-mL sterile screw-capped
plastic tube with O-ring. Leave the material dry.
- Scrape
the base of the vesicle or pustule with the blunt edge of the scalpel,
or with the wooden end of an applicator stick or swab.
- Smear
the scrapings onto a clean glass microscope slide.
- Apply
a microscope slide to the vesicular fluid multiple times, with progressive
movement of the slide, to make a touch prep.
- If a slide
is not available, swab the base of the lesion with a polyester or cotton
swab place in a screw-capped plastic vial, break off swab handle and
screw on lid. Do not add transport medium to the
vial.
- If available,
lightly touch “shiny side” of an electron microscope grid
to the unroofed base of the lesion. Repeat this procedure two more times,
varying the pressure applied to the unroofed lesion (lighter or firmer
pressure). Place in gridbox and record which slot is used for each patient
specimen.
- Allow
slides and grids to air dry for approximately 10 minutes.
- Store
slides in plastic slide holders for shipping. Parafilm may be used to
wrap the slide holder to prevent accidental opening. Store slides from
different patients in separate plastic slide
holder to prevent cross-contamination.
*For patients
with macular or papular rash, sample lesion via biopsy and acquire tonsillar
tissue swab, serum, and whole blood as described below.
Scab
lesion
- Sanitize
skin with an alcohol wide, allow to dry.
- Use a
26-gauge needle to pick/pry off as many scabs as possible (at least
four).
- Place
two scabs each in screw-capped plastic 1.5- to 2-mL vials.
- Use appropriate
sterile technique.
Lesion
biopsy
- Use appropriate
sterile technique and skin sanitation.
- Biopsy
lesion (2) with 3.5- or 4-mm biopsy punch (at least 2.5 mm; 3.5-4 mm
preferable).
- Place
one biopsy specimen in formalin.
- Place
one biopsy specimen in a 1.5- to 2-mL screw-capped vial. Do
not add any fluid.
Tonsillar
tissue swab
Swab or brush posterior tonsillar tissue and break off end of applicator
into a 1.5- to 2-mL screw-capped tube. Do not
add transport medium. Use polyester or Dacron swab.
Blood
samples
- Obtain
an acute-phase serum sample. Collect 7 to 10 cc of patient blood into
a marble-topped tube, or yellow-topped serum separator tube. Spin samples
to separate serum. Save the serum in at least two aliquots. Label tubes
as acute serum, with other information listed in Specimen Labeling (e.g.,
case ID number, date of collection). Store half of the serum at the
state health department and send half to CDC for testing.
- Obtain
whole blood sample. Collect 3 to 5 cc of blood into a purple-topped
tube. Gently mix blood with anticoagulant in tube to prevent clotting.
Label tube as whole blood, with other information listed in Specimen
Labeling (e.g., case ID number, date of collection). Send the tube with
whole blood to CDC for testing.
- Obtain
a convalescent-phase serum sample (4 to 6 weeks after collection of
acute-phase serum). Collect 7 to 10 cc of patient blood into a marble-topped
tube, or yellow-topped serum separator tube. Spin samples to separate
serum. Save the serum, and label the tube as convalescent serum, with
other information listed in Specimen Labeling (e.g., case ID number,
date of collection). Send convalescent-phase serum, together with the
remainder of the acute-phase sample stored at the health department,
to CDC for paired sera testing.
Note: For
pediatric patients, a minimum of 1 cc of whole blood is needed for testing.
If possible, collect at least 1 cc in each of an EDTA and clotting tube.
However, if only 1 cc can be obtained, use a clotting tube for collection.
V.
Stepwise Collection Procedure for Patients Identified 6-8 Weeks After
Onset of Rash Illness Suspected to be Monkeypox
Blood
samples
- Obtain
an initial serum sample when patient is first identified. Collect 7
to 10 cc of patient blood into a marble-topped tube, or yellow-topped
serum separator tube. Spin samples to separate serum. Save the serum
in at least two aliquots. Label tubes as acute serum, with other information
listed in Specimen Labeling (e.g., case ID number, date of collection).
Store half of the serum at the state health department and send half
to CDC for testing.
- Obtain
a follow-up serum sample 4 to 6 weeks after collection of initial serum
sample. Collect 7 to 10 cc of patient blood into a marble-topped tube,
or yellow-topped serum separator tube. Spin samples to separate serum.
Save the serum, and label the tube as convalescent serum with other
information listed in Specimen Labeling (e.g., case ID number, date
of collection). Send this second serum specimen, together with the remainder
of the initial sample stored at the health department, to CDC for paired
sera testing.
Note: For
pediatric patients, a minimum of 1 cc of whole blood is needed for testing.
If possible, collect 1 cc in both an EDTA and clotting tube. However,
if only 1 cc can be obtained, use a clotting tube for collection.
VI.
Stepwise Collection Procedures for Contacts of Persons with Monkeypox
Blood
samples
- Obtain
an initial serum sample when the person is first identified. Collect
7 to 10 cc of patient blood into a marble-topped tube, or yellow-topped
serum separator tube. Spin samples to separate serum. Save the serum
in at least two aliquots. Label tubes as acute serum, with other information
listed in Specimen Labeling (e.g., case ID number, date of collection).
Store half of the serum at the state health department and send half
to CDC for testing.
- Obtain
whole blood sample. Collect 3 to 5 cc of blood into a purple-topped
tube. Gently mix blood with anticoagulant in tube to prevent clotting.
Label tube as whole blood, with other information listed in Specimen
Labeling (e.g., case ID number, date of collection). Send the tube with
whole blood to CDC for testing.
- Obtain
a follow-up serum sample (4 to 6 weeks after collection of the initial
serum). Collect 7 to 10 cc of patient blood into a marble-topped tube,
or yellow-topped serum separator tube. Spin samples to separate serum.
Save the serum, and label the tube as convalescent serum, with other
information listed in Specimen Labeling (e.g., case ID number, date
of collection). Send convalescent-phase serum, together with the remainder
of the acute-phase sample stored at the health department, to CDC for
paired sera testing.
Note: For
pediatric patients, a minimum of 1 cc of whole blood is needed for testing.
If possible, collect at least 1 cc in each of an EDTA and clotting tube.
However, if only 1 cc can be obtained, use a clotting tube for collection.
VII.
Stepwise Collection Procedure for Patients Suspected to be in the Prodromal
Phase
- Tonsillar
tissue: Swab or brush posterior tonsillar tissue, then break off the
applicator tip into a 1.5- to 2-mL screw-capped tube. Do not add transport
medium. Use polyester or Dacron swab.
- Nasopharyngeal
swab: Swab nasopharynx to obtain nasopharyngeal secretions.
- Blood
samples.
- Obtain
an acute-phase serum sample. Collect 7 to 10 cc of patient blood
into a marble-topped tube, or yellow-topped serum separator tube.
Spin samples to separate serum. Save the serum in at least two aliquots.
Label tubes as acute serum, with other information listed in Specimen
Labeling (e.g., case ID number, date of collection). Store half
of the serum at the state health department and send half to CDC
for testing.
- Obtain
whole blood sample. Collect 3 to 5 cc of blood into a purple-topped
tube. Gently mix blood with anticoagulant in tube to prevent clotting.
Label tube as whole blood, with other information listed in Specimen
Labeling (e.g., case ID number, date of collection). Send the tube
with whole blood to CDC for testing.
- Obtain
a convalescent-phase serum sample (4 to 6 weeks after collection
of acute-phase serum). Collect 7 to 10 cc of patient blood into
a marble-topped tube, or yellow-topped serum separator tube. Spin
samples to separate serum. Save the serum, and label the tube as
convalescent serum, with other information listed in Specimen Labeling
(e.g., case ID number, date of collection). Send convalescent-phase
serum, together with the remainder of the acute-phase sample stored
at the health department, to CDC for paired sera testing.
VIII.
Post-collection Procedures
- After
specimen collection is completed, protective equipment worn by the specimen
collector should be removed. Disposable equipment (e.g., gown, gloves,
respirator, or mask) should be placed in a biohazard bag for disposal
with other medical waste. Reusable equipment (e.g., goggles, faceshield)
should be decontaminated and set aside for reprocessing. If cloth gowns
are used, they should be placed in a bag with other contaminated linen
in the patient's room.
- Needles
and other sharp instruments should be placed in a sharps container.
- Contaminated
waste generated through patient care should be handled in accordance
with existing facility procedures and local or state regulations for
regulated medical waste.
- Place
specimens from a single patient into a biohazard bag with an outside
label that includes:
- Patient
name
- Date
of collection
- Patient
date of birth.
- Package
specimens from a single patient (except biopsy specimens):
- On
gel packs at 4oC;
- Place
blood tubes (plastic or glass) in individual styrofoam holders;
- Place
all specimens in appropriate biosafety shipping containers in a
manner to withstand all shocks, pressure changes, or other conditions
incident to ordinary handling in transportation; and in a manner
to avoid leakage of contents.
- Package
non-formalin-fixed lesion biopsy specimens for shipping on dry ice,
leave formalin-fixed biopsy specimens at room temperature. Do not freeze
formalin-fixed biopsy sample.
- Specimens
may be stored in conditions outlined above if shipped within 24 hours
of collection. If this is not possible, store all samples except EM
grids, formalin-fixed tissue, and serum on dry ice or at –2oC
to –70oC until, and through, shipment. EM grids, formalin-fixed
tissue, and serum should be kept at 4oC until, and through
shipment. If there will be a delay in shipping, spin serum in marble
or yellow-top tubes to separate from clot, store at 4oC,
and ship at 4oC.
IX.
Shipping Diagnostic Specimens to CDC
- For information
about shipping specimens to CDC, see Instructions
for Packaging and Transport of Diagnostic Specimens for Monkeypox Laboratory
Testing.
- Label
the package as follows:
Centers
for Disease Control and Prevention
1600 Clifton Road, NE
ATTN: STAT Lab (forward to Poxvirus Section)
Atlanta, GA 30333
- Shipping
and specimen information should be submitted through CDC’s Web-based
incoming specimen tracking system by completing the form.
- Prior
to shipping the specimen, please send an e-mail to eoclogistic@cdc.gov
to alert CDC that the shipment is in transit.
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